Single hepatocyte labeling assay for evaluation of hepatocyte division. (A) Schematic representation of the labeling assay. Using the optimized HTVi in Rosa26-LacZ mice, single hepatocytes (singlets) are genetically labeled by low-frequency expression of LacZ. A singlet that underwent a cell division cycle would result in a pair of neighboring labeled hepatocytes (doublet), whereas an undivided singlet would remain as a singlet. The frequency of cell division is estimated by counting the numbers of singlets and doublets. (B) A liver section showing a singlet and a doublet. A section of a regenerated liver with labeled hepatocytes after 70% PHx is shown. The pair of labeled hepatocytes is a doublet (upper), and the labeled hepatocyte (lower) is an example of a singlet. Scale bar; 25 μm.